Zarnt T, Tradler T, Stoller G, Scholz C, Schmid F X, Fischer G
Forschungsstelle "Enzymologie der Proteinfaltung", Max-Planck-Gesellschaft, Kurt-Mothes-Str. 3, Halle/Saale, D-06120, Germany.
J Mol Biol. 1997 Sep 5;271(5):827-37. doi: 10.1006/jmbi.1997.1206.
The Escherichia coli trigger factor is a peptidyl-prolyl cis/trans isomerase (PPIase) which catalyzes proline-limited protein folding extremely well. It has been found associated with nascent protein chains as well as with the chaperone GroEL. The trigger factor utilizes protein regions outside the central catalytic domain for catalyzing refolding of unfolded proteins efficiently. Here we produced several fragments which encompass individual domains or combinations of the middle FKBP-like domain (M) with the N-terminal (N) and C-terminal (C) regions, respectively. These fragments appear to be stably folded. They show ordered structure and cooperative urea-induced unfolding transitions, and the far-UV CD spectrum of the intact trigger factor is well represented by the sum of the spectra of the fragments. This suggests that the native trigger factor shows a modular structure, which is composed of three fairly independent folding units. In the intact protein there is a slight mutual stabilization of these units. The high enzymatic activity in protein folding could not be restored by fusing alternatively the N or the C-terminal regions to the catalytic domain (in NM and MC constructs, respectively). Surprisingly, the high folding activity of the intact trigger factor has been regained partially by functional complementation of the overlapping NM and MC constructs.
大肠杆菌触发因子是一种肽基脯氨酰顺反异构酶（PPIase），能极其高效地催化脯氨酸受限的蛋白质折叠。已发现它与新生蛋白质链以及伴侣蛋白GroEL相关联。触发因子利用中央催化结构域之外的蛋白质区域来高效催化未折叠蛋白质的重折叠。在此，我们制备了几个片段，它们分别包含单个结构域，或者中间FKBP样结构域（M）与N端（N）和C端（C）区域的组合。这些片段似乎能稳定折叠。它们呈现出有序结构以及协同的尿素诱导的去折叠转变，完整触发因子的远紫外圆二色光谱很好地由这些片段光谱的总和表示。这表明天然触发因子呈现出模块化结构，由三个相当独立的折叠单元组成。在完整蛋白质中，这些单元之间存在轻微的相互稳定作用。通过分别将N端或C端区域与催化结构域融合（分别在NM和MC构建体中），无法恢复蛋白质折叠中的高酶活性。令人惊讶的是，通过重叠的NM和MC构建体的功能互补，部分恢复了完整触发因子的高折叠活性。
